EC50 Concentration Research Articles

Impact of geology and anthropogenic activities over the water quality with emphasis on fluoride in water scarce Lalitpur district of Bundelkhand region, India

The Bundelkhand region of India is suffering from acute water scarcity, raising concern over the potability in the region. Therefore, to develop a baseline data set of groundwater quality, sampling was carried out from the 110 existing shallow hand pumps and tube wells covering the Lalitpur district. Groundwater samples were investigated for hydro-geochemical and isotopic signatures (δ18O and δ2H) to understand the driving factors leading to water quality and its contamination in the region. The results of Hierarchical cluster analysis revealed four different clusters according to their water quality. Cluster 1 and 2 water samples have a good quality of water and these samples fall in the vicinity of major or minor drainage networks of the area. Whereas, clusters 3 and 4 are of deteriorated water quality and located far-off from the drainage networks in the study area. The findings from chemical analysis and chemometric method suggest that the groundwater composition is mainly influenced by rock weathering and anthropogenic activities. Fluoride exposure dosage for the infant and children is twice that of adults in the study area, indicating a stronger impact of fluoride concentration in infants and children. The stable isotopic analysis shows that origin of groundwater is local precipitation, with evaporative enrichment in groundwater. The groundwater of cluster 3 and 4 shows evaporative nature along with high EC and Cl concentration. The variation of concentration of ions in the study area prevails along the groundwater flow direction and surface drainage reveals the control of hydrogeological attributes in the groundwater.

Characterization of Potency of the P2Y13 Receptor Agonists: A Meta-Analysis.

P2Y13 is an ADP-stimulated G-protein coupled receptor implicated in many physiological processes, including neurotransmission, metabolism, pain, and bone homeostasis. Quantitative understanding of P2Y13 activation dynamics is important for translational studies. We systematically identified PubMed annotated studies that characterized concentration-dependence of P2Y13 responses to natural and synthetic agonists. Since the comparison of the efficacy (maximum response) is difficult for studies performed in different systems, we normalized the data and conducted a meta-analysis of EC50 (concentration at half-maximum response) and Hill coefficient (slope) of P2Y13-mediated responses to different agonists. For signaling events induced by heterologously expressed P2Y13, EC50 of ADP-like agonists was 17.2 nM (95% CI: 7.7–38.5), with Hills coefficient of 4.4 (95% CI: 3.3–5.4), while ATP-like agonists had EC50 of 0.45 μM (95% CI: 0.06–3.15). For functional responses of endogenously expressed P2Y13, EC50 of ADP-like agonists was 1.76 μM (95% CI: 0.3–10.06). The EC50 of ADP-like agonists was lower for the brain P2Y13 than the blood P2Y13. ADP-like agonists were also more potent for human P2Y13 compared to rodent P2Y13. Thus, P2Y13 appears to be the most ADP-sensitive receptor characterized to date. The detailed understanding of tissue- and species-related differences in the P2Y13 response to ADP will improve the selectivity and specificity of future pharmacological compounds.

Abstract P750: 3K3A-APC Restores Oligodendrocyte Pools in Models of White Matter Stroke via PAR1 Signaling

A recombinant variant of activated protein C, 3K3A-APC, was recently tested in a successful phase II clinical trial for ischemic stroke (RHAPSODY), which demonstrated that the treatment may reduce hemorrhages and is safe in patients that were treated with tPA and/or thrombectomy. Animal models of large ischemic stroke (middle cerebral artery occlusion) and neurodegeneration have shown that the effects of 3K3A-APC are mediated by preserved APC cell signaling activities (anti-apoptotic neuronal protection, protection of blood-brain barrier (BBB) integrity and anti-inflammatory activities). Because recent studies have suggested a role of protease activated receptor 1 (PAR1) in oligodendrocyte recovery after white matter damage, we wanted to test whether biased PAR1 signaling mediated by 3K3A-APC would protect oligodendrocytes in in vivo and ex vivo models of ischemic white matter stroke. First, we used a mouse model of white matter stroke induced by injecting a vasoconstrictor N 5 -(1-Iminoethyl)-L-ornithine (L-NIO) into the corpus callosum. Our results show that treatment with recombinant murine 3K3A-APC (0.2 mg/kg, starting 4h after stroke) protected oligodendrocytes (OLS, Olig2+CNPase+) and oligodendrocyte precursor cells (OPCs, Olig2+PDGFRα+) from cell death at 1 and 7 days after stroke. In vitro , 3K3A-APC protected primary mouse oligodendrocytes from apoptosis when exposed to OGD conditions at nanomolar concentrations (EC50 concentration was 5.2 nM). Improved oligodendrocyte viability in vivo was associated by about ~67% reduction in overall cell death (cresyl violet staining, T2w and diffusion tensor MRI) and reduced BBB leakage (fibrin staining and DCE-MRI) as well as improved functional recovery (adhesive removal and grid walking tests). Protective effects of 3K3A-APC were abolished by silencing of PAR1 with small interfering RNA (siRNA) in vivo and in vitro . In conclusion, PAR1 mediated signaling initiated by 3K3A-APC protects oligodendrocyte pools in models of ischemic white matter damage with preservation of neurovascular unit integrity.

On the Applicability of Chitosan Oligomers-Amino Acid Conjugate Complexes as Eco-Friendly Fungicides against Grapevine Trunk Pathogens

In a context in which the incidence and severity of grapevine fungal diseases is increasing as a result of both climate change and modern management culture practices, reducing the excessive use of phytosanitary products in viticulture represents a major challenge. Specifically, grapevine trunk diseases (GTDs), caused by several complexes of wood decay or xylem-inhabiting fungi, pose a major challenge to vineyard sustainability. In this study, the efficacy of chitosan oligomers (COS)–amino acid conjugate complexes against three fungal species belonging to the Botryosphaeriaceae family (Neofusicoccum parvum, Diplodia seriata, and Botryosphaeria dothidea) was investigated both in vitro and in planta. In vitro tests led to EC50 and EC90 effective concentrations in the 254.6−448.5 and 672.1−1498.5 µg·mL−1 range, respectively, depending on the amino acid involved in the conjugate complex (viz. cysteine, glycine, proline or tyrosine) and on the pathogen assayed. A synergistic effect between COS and the amino acids was observed against D. seriata and B. dothidea (synergy factors of up to 2.5 and 2.8, respectively, according to Wadley’s method). The formulations based on COS and on the conjugate complex that showed the best inhibition rates, COS−tyrosine, were further investigated in a greenhouse trial on grafted vines of two varieties (”Tempranillo” on 775P and “Garnacha” on 110R rootstock), artificially inoculated with the mentioned three Botryosphaeriaceae species. The in planta bioassay revealed that the chosen formulations induced a significant decrease in disease severity against N. parvum and B. dothidea. In summary, the reported conjugate complexes may be promising enough to be worthy of additional examination in larger field trials.

The endemic halophyte Sarcocornia carinata Fuente, Rufo & Sánchez-Mata (Chenopodiaceae) in relation to environmental variables: elemental composition and biominerals

AimsWe propose a thorough study of the succulent halophyte Sarcocornia carinata endemic to the saline lagoons of the center of the Iberian Peninsula. We describe its elemental composition and possible seasonal variation in relation to edaphic and climatic variables, identify biominerals and analyze the distribution of salt ions and biominerals in tissue.MethodsPlants and edaphic samples were collected in the four seasons of 1 year. Soils were analyzed for their pH, EC, color, and bioavailable concentration of Na+, K+, Mg2+, Ca2+, Cl−, SO42−. Soils and plants were analyzed for their total elemental and mineralogical composition. The distribution of elements and minerals in tissues was studied by scanning electron microscopy.ResultsDespite the variations observed in the edaphic and climatic variables, the variables studied in the plants varied slightly throughout the year. In the plants, Mg was the element that reflected climatic changes the most, while the K and Ca concentrations did not vary. Salty precipitates and crystallizations were distributed mainly in the epidermis, water storage parenchyma, cortex, and vascular vessels. Several crystals observed were compatible with halite, gypsum, glushinskite and weddellite.ConclusionsThe study corroborates that inland S. carinata behaves in the same way as other littoral succulent euhalophytes and reinforces the hypothesis that the concentration of elements and quantitative abundance pattern depend largely on the main adaptation mechanisms of halophytes.

Azolla (Caroliniana): An Aquatic Energy Crop for Remediation of Eutrophic Ecosystems with Prospect of Biofuel Production in Bangladesh

The Phytoremediation ability of aquatic macrophyte, Azolla as potential biomass, to improve the quality of eutrophic lakes and river water of Bangladesh at lab scale were studied to remove organics, solutes and improving the physicochemical properties of water including pH, DO, EC, TDS, Turbidity and NaCl concentration. In the different eutrophic water, a 100% survival rate of the species was observed. The extensive root system of the phytoremediator improves water quality after one week as determined by a marked decrease in Turbidity, TDS, EC & Nacl and increased in DO and pH. DO content served as a useful indicator of water quality and increased manifolds over 7 days (one week) of treatment. The capability of the plant in removing pollutants and improving water quality was established from the study. After phytoremediation activities, the discarded energy crop biomass could be used to produce green energy because of its high growth potential.

Isolation and Antitrypanosomal Characterization of Furoquinoline and Oxylipin from Zanthoxylum zanthoxyloides.

In the absence of vaccines, there is a need for alternative sources of effective chemotherapy for African trypanosomiasis (AT). The increasing rate of resistance and toxicity of commercially available antitrypanosomal drugs also necessitates an investigation into the mode of action of new antitrypanosomals for AT. In this study, furoquinoline 4, 7, 8-trimethoxyfuro (2, 3-b) quinoline (compound 1) and oxylipin 9-oxo-10, 12-octadecadienoic acid (compound 2) were isolated from the plant species Zanthoxylum zanthoxyloides (Lam) Zepern and Timler (root), and their in vitro efficacy and mechanisms of action investigated in Trypanosoma brucei (T. brucei), the species responsible for AT. Both compounds resulted in a selectively significant growth inhibition of T. brucei (compound 1, half-maximal effective concentration EC50 = 1.7 μM, selectivity indices SI = 74.9; compound 2, EC50 = 1.2 μM, SI = 107.3). With regards to effect on the cell cycle phases of T. brucei, only compound 1 significantly arrested the second growth-mitotic (G2-M) phase progression even though G2-M and DNA replication (S) phase arrest resulted in the overall reduction of T. brucei cells in G0-G1 for both compounds. Moreover, both compounds resulted in the aggregation and distortion of the elongated slender morphology of T. brucei. Analysis of antioxidant potential revealed that at their minimum and maximum concentrations, the compounds exhibited significant oxidative activities in T. brucei (compound 1, 22.7 μM Trolox equivalent (TE), 221.2 μM TE; compound 2, 15.0 μM TE, 297.7 μM TE). Analysis of growth kinetics also showed that compound 1 exhibited a relatively consistent growth inhibition of T. brucei at different concentrations as compared to compound 2. The results suggest that compounds 1 and 2 are promising antitrypanosomals with the potential for further development into novel AT chemotherapy.

THER-04. IS THERE A ROLE FOR CANNABIDIOL IN THE TREATMENT OF CHILDHOOD BRAIN TUMOURS?

Brain tumours are the leading cause of cancer related death in children with limited treatment options and high recurrence rates. Recent evidence suggests there may be anti-tumoral properties of cannabinoids, and of cannabidiol (CBD) in particular. We evaluated the effect of CBD on paediatric brain tumour cell lines in 2D and 3D spheroids; pHGG (SF188), ependymoma (BxD-1425EPN) and human astrocytes. At the CBD EC50 concentration, astrocytic cell death was insignificant. 3D spheroids decreased in size by approximately 20% when cultured in CBD compared to cells only after 5-day exposure. Cell death increased with time after a single dose of CBD. Western Blot showed an increase in Lc3b expression (autophagy) after 24 hours incubation (early cell death) with CBD in both BxD-1425EPN and SF188 with PARP expression (apoptosis) increased after 5 days incubation (late cell death). Cell cycle analysis showed a decrease of cells in G1 and no change in G2 indicating cell cycle arrest. In hypoxia, SF188 and BxD-1425EPN cells showed decreased cell death after 24 hours and 5 days when compared to normoxia and an EC50 within acceptable limits could not be achieved. SF188 cells pre-treated with receptor antagonists indicate that CBD was not acting through CB1, CB2, GPR18, PPARα or PPARγ receptors but may act as a partial agonist of the TRPV1 and 5-HT1A receptors and a full agonist of the GPR55 receptor (resazurin assay). This provides evidence that CBD is effective at killing paediatric brain tumour cells and does not have a significant effect on normal astrocytes.

Comparative Study of Lipophilic Statin Activity in 2D and 3Din vitro Models of Human Breast Cancer Cell Lines MDA-MB-231 and MCF-7.

IntroductionStatins are a type of drugs that are used to lower cholesterol level in blood. Since the early 1990s, it has been known that statins could be beneficial in cancer therapy. However, data remain controversial, especially regarding estrogen receptors status. Despite many studies in breast cancer models in vitro, the correlations of effects of separate statins in various model systems remain unclear.AimOur aim was to evaluate the anticancer activity of lovastatin, mevastatin, pitavastatin and simvastatin on different subtypes of human breast cancer (MDA-MB-231 and MCF-7 cell lines) in spatially different 2D and 3D cultures in vitro.Materials and MethodsCell viability was tested using MTT assay. Effect of statins on cell colony formation was evaluated by calculating breast cancer cell colony area and number. The effect on cell migration was estimated by “wound healing” assay. The activity of compounds in 3D cultures was evaluated by measuring the spheroid size changes during incubation.ResultsAmong the tested statins, pitavastatin had the greatest effect on the viability of breast cancer MDA-MB-231 and MCF-7 cell lines. The mevastatin and pitavastatin mostly decreased the MDA-MB-231 and MCF-7 cell colony formation. All statins at 90% of their estimated effective concentration (EC50) and simvastatin at 10% of its EC50 concentration suppressed the MCF-7 cells migration in “wound healing” assay. Only higher concentrations of mevastatin and pitavastatin slowed down the MDA-MB-231 cell migration. Statins showed different activity on 3D cell cultures growth. Lovastatin and simvastatin delayed the growth of MDA-MB-231 cell spheroids, while mevastatin and pitavastatin reduced the growth of MCF-7 spheroids.ConclusionStatins possess different anticancer activity in human breast cancer MDA-MB-231 and MCF-7 cell cultures. Pitavastatin and simvastatin showed the highest activity in most tested assays, especially against MCF-7 cell line.

Similar toxicity mechanisms between graphene oxide and oxidized multi-walled carbon nanotubes in Microcystis aeruginosa

In photosynthetic microorganisms, the toxicity of carbon nanomaterials (CNMs) is typically characterized by a decrease in growth, viability, photosynthesis, as well as the induction of oxidative stress. However, it is currently unclear how the shape of the carbon structure in CNMs, such as in the 1-dimensional carbon nanotubes (CNTs) compared to the two-dimensional graphene oxide (GO), affects the way they interact with cells. In this study, the effects of GO and oxidized multi-walled CNTs were compared in the cyanobacterium Microcystis aeruginosa to determine the similarities or differences in how the two CNMs interact with and induce toxicity to cyanobacteria. Using change in Chlorophyll a concentrations, the effective concentrations inducing 50% inhibition (EC50) at 96 h are found to be 11.1 μg/mL and 7.38 μg/mL for GO and CNTs, respectively. The EC50 of the two CNMs were not found to be statistically different. Changes in fluorescein diacetate and 2′,7′-dichlorodihydrofluorescein diacetate fluorescence, measured at the EC50 concentrations, suggest a decrease in esterase enzyme activity but no oxidative stress. Scanning and transmission electron microscopy imaging did not show extensive membrane damage in cells exposed to GO or CNTs. Altogether, the decrease in metabolic activity and photosynthetic activity without oxidative stress or membrane damage support the hypothesis that both GO and CNTs induced indirect toxicity through physical mechanisms associated with light shading and cell aggregation. This indirect toxicity explains why the intrinsic differences in shape, size, and surface properties between CNTs and GO did not result in differences in how they induce toxicity to cyanobacteria.

Polyphenols Content and Antimicrobial, Antioxidant and Hemolytic Activities of Essential Oils from Four Selected Medicinal Plants Growing in Algeria

The Saharan and steppe spontaneous plants are very characteristic because of their particular adaptation to the desert and extreme environment. Some species have pharmacological properties that give them a medicinal interest. The aim of the present work was to determine the polyphenol contents of essential oils obtained from four endemic plants growing in Algeria (Pituranthos scoparius, Myrtus nivellei, Rosmarinus officinalis and Mentha piperita), and study its biological activity, including antimicrobial, antioxidant, and hemolytic. The antimicrobial activity was evaluated by the microdilution method against twelve strains. The antioxidant activity was carried out by two methods (DPPH radical scavenging and reducing power). However, the hemolytic effect has been evaluated against the red blood cells. P. scoparius and M. piperita showed yields of essential oils higher than 1%. All the strains showed sensitivity against the essential oils tested with the exception of the C. albicans treated by R. officinalis essential oils. The most sensitive strain was C. albicans treated by P. scoparius essential oils by MIC of 0.0781 mg/mL, it was the same plant that shows the highest polyphenol content (14.78 ± 0.72 g GAE/g DS). The antioxidant activity by the DPPH method was greater for all essential oils tested by IC50 ranging from 0.69 ± 0.07 (R. officinalis) to 30.67 ± 2.12 mg/mL (M. nivellei). The R. officinalis essential oils reported more antioxidant power than the positive control (ascorbic acid). In reducing iron, it was the R. officinalis essential oils which were found to be the most active with an EC50 concentration of 9.67 ± 1.36 mg/mL. After 120 min incubation, minimal haemolysis (10%) was obtained with essential oils of R. officinalis at a concentration of 0.39 mg/mL. We conclude that P. scoparius essential oils showed the high content of polyphenols and R. officinalis essential oils reported more antioxidant power than the positive control (ascorbic acid).

Preclinical Investigation of the p53-MDM2 Antagonist Idasanutlin (RG7388) Demonstrates Significant Activity in High Risk Adult Acute Lymphoblastic Leukemia

Outcomes for adult patients with acute lymphoblastic leukemia (ALL) are dismal and have not kept pace with their pediatric counterparts, with five-year survival rates of less than 45%. TP53 mutations are infrequent in ALL, but activity of the oncoprotein MDM2 may otherwise phenotypically disrupt and circumvent normal p53 function, positing the p53-MDM2 signaling axis as a potential therapeutic target for the engagement of intrinsic cell death programs. Given the clinical safety and responses to p53-MDM2 antagonist idasanutlin (RG7388) in other hematological cancers and solid tumors, we aimed to evaluate the therapeutic potential of idasanutlin in ALL. Single-agent activity of idasanutlin was investigated in 17 high-risk ALL patient and patient-derived xenograft samples (aged 4 to 51 years), both B-ALL (n=15) and T-ALL (n=2), including KMT2A-rearranged, TCF3-rearranged, and Philadelphia-positive ALL. An ex vivo coculture of ALL blasts and hTERT-immortalized mesenchymal stem cells (MSC) was employed to support growth of the ALL blasts during short-term culture, complemented by a fluorescent image-based microscopy platform which identifies and discriminates the two cellular compartments using random forest machine learning algorithms based on cellular nuclear staining. Idasanutlin demonstrated sub-micromolar, dose-dependent anti-leukemic activity against 15 of 17 samples tested, with half maximal effective concentrations (EC50) in the range of 10 to 220 nM (mean EC50 = 45.1nM); the two exceptions were later determined to harbor homozygous inactivating TP53 mutations; p.Y220C and p.S241P within the p53 DNA binding domain. The idasanutlin EC50 concentrations determined are clinically achievable, well below the peak plasma concentrations reported in patients for other disease indications. Furthermore, idasanutlin concentrations below 10µM had no impact upon MSC survival. Conforming to p53-MDM2 auto-regulatory feedback mechanisms, we demonstrated that idasanutlin efficiently stabilized and activated p53 at the protein level within 6 hours when treated with their respective idasanutlin EC50, to a level greater than 4-fold increased relative to their respective vehicle-only controls (p=0.001, n=7). Further, p53-regulated transcriptional target gene products, MDM2 and p21, were increased by 5-fold and 2-fold respectively, validating engagement of the p53 pathway by idasanutlin (p=0.036 and 0.125, respectively). By contrast, idasanutlin did not elicit increased expression of either p53-regulated transcriptional target gene product in the identified TP53-mutant patient samples (n=2). On-target specificity of idasanutlin was further confirmed in a NALM6 isogenic cell line model, whereby the TP53 wildtype line was sensitive to idasanutlin (EC50 = 74nM) and effectively activated p53 signaling whereas the TP53 homozygous null line was highly resistant (EC50 = ~10µM). To determine whether the decreased cell numbers and engagement of p53 signaling observed were accompanied by cell death, the capacity of idasanutlin to induce apoptosis in the ALL samples was next investigated. Patient-derived ALL samples (n=6) were treated with vehicle or idasanutlin at their respective EC50s for 24 and 48 hours, and then analyzed by flow cytometry. There was an increase in annexin-V positive cells within 24 hours compared to the vehicle-only treated cells (mean±SD 14.3±6.6% vs 27.0±21.2% respectively (p=0.125). By 48 hours significant apoptosis was attained, with a mean±SD of 59.0±23.8% annexin-V positive cells compared to the mean of vehicle-treated cells at 29.1±11.6% (p=0.004). Cleaved poly(ADP-ribose) polymerase (PARP) levels were also increased greater than 3-fold compared to vehicle-only control cells as assessed by immunoblotting (p=0.045, n=3), corroborating these findings. These data emphasize the potential of pharmacologically targeting the p53-MDM2 axis in ALL, demonstrating potent, on-target, cytotoxic activity in a range of high-risk ALL cytogenetic subgroups. Taken together, these findings support further preclinical investigations into idasanutlin and other p53-MDM2 antagonists and potential combinations to improve the treatment of adult ALL. Disclosures Irving: F. Hoffmann-La Roche: Research Funding.

Protective effect of Catharanthus roseus plant extracts against endosulfan and its isomers induced impacts on non-targeted insect model, Drosophila melanogaster and live brain cell imaging

Endosulfan has been recognized as a highly controversial pesticide due to its acute toxicity, potential bioaccumulation, persistency, and long-range atmospheric transport. Several plant extracts act as antioxidant agents against wide-range of pesticide toxicity hazards through the free radicals scavenging properties. Plants' secondary metabolites are considered as efficient protective agents against various cellular toxic injuries. Understanding these properties of botanicals, several researchers currently focused on the detoxification and ameliorative potency of plant extracts against highly toxic chemicals. In our studies, we focused on the endosulfan total and its isomers (alpha and beta) induced changes on Drosophila melanogaster and their ameliorative effects by co-administrated with methanolic and aqueous extracts of Catharanthus roseus whole plant. We selected the 1/5th EC50 concentration of alpha-endosulfan, beta-endosulfan, and endosulfan (total) and co-administrated with 1/50th EC50 concentration of aqueous and methanolic extracts and evaluated their ameliorative effects, in terms of verifying the life stage activities, protein profiling and also by using live brain cells imaging. We finally concluded that, the methanolic and aqueous extracts inhibit the toxic impacts caused by endosulfan and its isomers and also increasing the survival rate of the test organism.

Making the Biotic Ligand Model kinetic, easier to develop, and more flexible for deriving water quality criteria

In aquatic environments, the ecological risks posed by metals are greatly affected by water chemistry, thereby creating challenges for water quality management. Biotic ligand models (BLMs) have become the most widely used tools to interpret and predict water chemistry effects. Traditional BLM development methods require a large number of toxicity tests and organisms, and model predictions are limited to certain toxicity statistics (e.g., 48-h median effective concentration, 48-h EC50), to which the models were calibrated. To address these limitations, we propose a new method to develop BLMs by integrating them into the toxicokinetic-toxicodynamic (TK-TD) framework. Metal bioaccumulation was predicted from metal exposure and water chemistry using the BLM-type toxicokinetics, whilst metal toxicity was predicted from metal bioaccumulation using the toxicodynamics. Using the new method, we developed a kinetic BLM of cadmium for Daphnia magna with only six toxicity tests and 1540 daphnids; this represents a 60−80% reduction compared to the traditional methods. The model was validated in the presence of commercial dissolved organic matter (DOM) and in natural waters sampled from 12 lakes. The kinetic BLM was able to accurately simulate the protective effects of the commercial DOM by employing the Stockholm humic model, whilst the complexation capabilities of some natural DOM were overestimated. We further used the model to predict Cd EC50 and no-effect concentrations for different waters, generating predictions close to the effect concentrations reported in the literature. Overall, our method requires fewer resources and presents an easier approach to develop BLMs; the kinetic BLM is more flexible and can serve as a useful tool for developing water quality criteria.

Antifungal Activity of Chitosan Oligomers–Amino Acid Conjugate Complexes against Fusarium culmorum in Spelt (Triticum spelta L.)

Fusarium head blight (FHB) is a complex disease of cereals caused by Fusarium species, which causes severe damages in terms of yield quality and quantity worldwide, and which produces mycotoxin contamination, posing a serious threat to public health. In the study presented herein, the antifungal activity against Fusarium culmorum of chitosan oligomers (COS)–amino acid conjugate complexes was investigated both in vitro and in vivo. The amino acids assayed were cysteine, glycine, proline and tyrosine. In vitro tests showed an enhancement of mycelial growth inhibition, with EC50 and EC90 effective concentration values ranging from 320 to 948 µg·mL−1 and from 1107 to 1407 µg·mL−1 respectively, for the conjugate complexes, as a result of the synergistic behavior between COS and the amino acids, tentatively ascribed to enhanced cell membrane damage originating from lipid peroxidation. Tests on colonies showed a maximum percentage reduction in the number of colonies at 1500 µg·mL−1 concentration, while grain tests were found to inhibit fungal growth, reducing deoxynivalenol content by 89%. The formulation that showed the best performance, i.e., the conjugate complex based on COS and tyrosine, was further investigated in a small-scale field trial with artificially inoculated spelt (Triticum spelta L.), and as a seed treatment to inhibit fungal growth in spelt seedlings. The field experiment showed that the chosen formulation induced a decrease in disease severity, with a control efficacy of 83.5%, while the seed tests showed that the treatment did not affect the percentage of germination and resulted in a lower incidence of root rot caused by the pathogen, albeit with a lower control efficacy (50%). Consequently, the reported conjugate complexes hold enough promise for crop protection applications to deserve further examination in larger field trials, with other Fusarium spp. pathogens and/or Triticum species.

Growth inhibition associated with disruption of the actin cytoskeleton by Latrunculin A in rhabdomyosarcoma cells.

Functional genomic screening of KRAS-driven mouse sarcomas was previously employed to identify proliferation-relevant genes. Genes identified included Ubiquitin-conjugating enzyme E2 (Ube2c), Centromere Protein E (Cenpe), Hyaluronan Synthase 2 (Has2), and CAMP Responsive Element Binding Protein 3 Like 2 (Creb3l2). This study examines the expression and chemical inhibition of these candidate genes, identifying variable levels of protein expression and significant contributions to rhabdomyosarcoma (RMS) cell proliferation. Chemical treatment of human and murine RMS cell lines with bortezomib, UA62784, latrunculin A and sorafenib inhibited growth with approximate EC50 concentrations of 15-30nM for bortezomib, 25-80nM for UA62784 and 80-220nM for latrunculin A. The multi-kinase inhibitor sorafenib increased in vitro proliferation of 4 of 6 sarcoma cell lines tested. Latrunculin A was further associated with disruption of the actin cytoskeleton and reduced ERK1/2 phosphorylation. Together, this work advances opportunities for developing therapies to block progression of soft-tissue sarcomas and demonstrates that disruption of the actin cytoskeleton in sarcoma cells by latrunculin A is associated with a reduction in RMS cell growth. (167 words).

Opuntia ficus-indica cladode extract is an embryotoxic, larvicidal, and oviposition-deterrent agent for the diamondback moth, Plutella xylostella

The search for new compounds to control Plutella xylostella, the main pest of Brassica crops, has been stimulated by the environmental impact of the insecticides currently used and reports of resistant populations. This work evaluated the effects of a cladode extract from Opuntia ficus-indica (CE) on the survival and development of P. xylostella, as well as on its oviposition behavior. Additionally, the effects on the histological organization of the larval midgut, as well as digestive enzyme activities, were evaluated. CE contained flavonoids, cinnamic derivatives, terpenes, steroids, and reducing sugars. High-performance liquid chromatography analysis confirmed the presence of rutin and compounds containing aliphatic chains and aromatic rings in their structures. CE reduced the viability of eggs with an EC50 (concentration that reduces egg viability by 50%) at 5.37% (w/v) and killed larvae with an LC50 (concentration that kills 50% of larvae) at 7.75% (w/v). Histological analysis revealed several alterations in the midgut of larvae exposed to the extract, including the presence of spherites, cell nuclei expulsion, the disappearance of the peritrophic membrane, and an increase in the number of goblet cells. CE inhibited the in vitro activity of proteases from larvae midgut extracts. The extract also presented oviposition-deterrent activity. In conclusion, cladodes of O. ficus-indica are a source of compounds toxic to eggs and larvae of P. xylostella and are able to interfere with oviposition site choice. The results suggest potential of CE as a botanical insecticide, though field trials are needed to confirm activity.

Essential oils against Varroa destructor: a soft way to fight the parasitic mite of Apis mellifera

Essential oils (EOs) extracted from the aromatic plants Artemisia annua, Artemisia verlotiorum, Cinnamomum verum, and Citrus reticulata were investigated as repellents against the honey bees parasitic mite Varroa destructor. In laboratory tests, all EOs except C. reticulata exerted significant repellent activity against the mite after 24 h exposure. C. verumwas the most effective EO against V. destructor (median effective concentration EC50 =1.30 μL L−1), and the least toxic against honey bees (EC50 = 13.29 μL L−1). Because of its high selectivity ratio (A. mellifera LD50/V. destructor EC50 = 10.22), C. verum EO was then tested to control varroosis in colonies in field trials. The results of open field tests confirmed the efficacy observed in the laboratory. After one week of treatment, colonies treated with the EO showed a significant reduction of V. destructor infestation (about 65% at the dose of 25.0 μL L−1) and no negative effects on A. mellifera. Overall, our experiment indicated that C. verum EO could be used to effectively control varroosis in the hive with no side effects on the bee colonies.

In Vitro Assessment of the Antioxidant and Antitumor Potentials of Biogenic Silver Nanoparticle

Silver nanoparticles (AgNPs) were biosynthesized using the cell free supernatant of putative probiotic Lactobacillus paracasei A26. Several biological activities of biogenic AgNPs were investigated in respect to in vitro anti-oxidant and anti-tumor potentials. Anti-oxidant potentials were screened in terms of free radical scavenging activity against two free radicals, 2, 2-Diphenyl-1-picrylhydrazyl (DPPH) and resazurin dye. AgNPs exhibited a potent scavenging activity against resazurin dye (91±0.046%) with an EC50 concentration of 146.823 µg/ml, while scavenging of DPPH was significantly (P≤0.05) reduced to 72.330±0.114% using a higher EC50 concentration of 176.12 µg/ml. The anti-tumor potentials of biogenic AgNPs were studied in relation to the cytotoxicity against two human breast cancer cell lines (CAL-51 and MCF7), using crystal violet dye assay. The viability of AgNPs-treated cancerous cells was significantly decreased in a time- and concentration manner, as compared to insignificant cytotoxic effects against the normal cell line. However, the anti-proliferative activity of AgNPs did not exceed the value of 63.85±0.019% in both cancer cell lines. CAL-51 cells were the most sensitive to the introduced AgNPs, with a maximum decrease in viability of 49.889±0.021% being reached using an IC50 value of 98.65µg/ml for 48h exposure time. The inhibition percentage was increased to 60.13±0.005% when the used IC50 value was significantly declined to 40.73µg/ml with an exposure time expanded to 72h. MCF7 cells showed lower sensitivity than CAL-51 cells, but with a similar inhibition trend of 59.523±0.01% with an IC50 concentration of 66.54 µg/ml for 48h which was increased to 63.857±0.019% when the IC50 was reduced to 62.63 µg/ml and the exposure time expanded to 72h. The morphological changes of AgNPs-treated cells were apparent at 72h exposure time, with cells showing apoptotic-like features such as shrinkage and losing of regular fusiform shape. Moreover, cells became detached to surfaces and from each other.

ROR1 Is Expressed in Diffuse Large B-Cell Lymphoma (DLBCL) and a Small Molecule Inhibitor of ROR1 (KAN0441571C) Induced Apoptosis of Lymphoma Cells.

The receptor tyrosine kinase ROR1 is absent in most normal adult tissues, but overexpressed in several malignancies. In this study, we explored clinical and functional inhibitory aspects of ROR1 in diffuse large B-cell lymphoma (DLBCL). ROR1 expression in tumor cells was more often observed in primary refractory DLBCL, Richter’s syndrome and transformed follicular lymphoma than in relapsed and non-relapsed DLBCL patients (p < 0.001). A survival effect of ROR1 expression was preliminarily observed in relapsed/refractory patients independent of gender and stage but not of age, cell of origin and international prognostic index. A second generation small molecule ROR1 inhibitor (KAN0441571C) induced apoptosis of ROR1+ DLBCL cell lines, similar to venetoclax (BCL-2 inhibitor) but superior to ibrutinib (BTK inhibitor). The combination of KAN0441571C and venetoclax at EC50 concentrations induced almost complete killing of DLBCL cell lines. Apoptosis was accompanied by the downregulation of BCL-2 and MCL-1 and confirmed by the cleavage of PARP and caspases 3, 8, 9. PI3Kδ/AKT/mTOR (non-canonical Wnt pathway) as well as β-catenin and CK1δ (canonical pathway) were inactivated. In zebra fishes transplanted with a ROR1+ DLBCL cell line, KAN0441571C induced a significant tumor reduction. New drugs with mechanisms of action other than those available for DLBCL are warranted. ROR1 inhibitors might represent a novel promising approach.